Clinical Pathology

276 Pages·2012·7.9 MB·English

COMSTOCK SERIES IN VETERIN AR Y MEDICINE

WILLIAM ARTHUR HAGAN, D V.M., D.se., Consulting Edtfor

MANUAL OF Veterinary

Clinical Pathology

Manual of Veterinary

Clinical Pathology

REVISED AND AMPLIFIED

BY David L. Coffin, v. M. D. * SCHOOL OF
,
..

VETERINARY MEDICINE, UNIVERSITY OF PENNSYLVANIA

ITHACA, NEW YORK * 1945

Comstock Publishing Com pany, Inc

COPYRIGHT 1945

BY COMSTOCK PUBLISHING COMPANY, INC.

All rtghts rc,crvcd ThIs book, or
parts thereof, must 110t be re-
produced In Jny fOlm wlthom
pertms,ion in \, fltIng b om the
publ "hu, except hy .1 rCI'lcwer
who wIshes to quote brIef pas·

saJl"~s In a revIew of the book.

PRiNTFD
IN
TllF
UNITFD
STATE~

OF AMlRlrA BY 'THI:
VA)L nAl LOU

PRFSS~ INC ••
UI"fGH'\MfON.
N
Y.

PREFACE

T
HIS BOOK has been wri.tten to fill the need for a handy and concise
saurce of information on the most practical laboratory procedures
now in use for the diagnosis of animal disease. It has been organized to
serve the practicing veterinarian as well as the veterinary student. It is
hoped that this treatment of the subject will also prove useful to labora-
tory diagnosticians, veterinary pathologists and research workers in ani-
mal problems.
The first edition of the Manual was an enlargement of the laboratory
notes used in the combined course of Clinical Pathology and Laboratory
Diagnosis at the University of Pennsylvania, School of Veterinary Medi-
cine. This new edition is a revision of the original edition published by
the author in 1944. New material has been selected to make the Manual a
freshly sharpened tOG!.
'. ,;
Practical, every-day techniques are given in ordered steps with the hope
that they m:ly be fGllowed without confusion by persons not especially
experienced 1Il laboratory work. Field methods, the procedures for col-
lecting specimens from field cases and the best methods of preserving
such specimens are considered. Special techniques, having more value to
those engaged in research or in laboratory diagnosis, are set in smaller
type. These may be omitted by the student or the practitioner without
disturbing the continuity of the text. Interpretation of laboratory findings
is stressed because the de~ired end result of any laboratory examination
is a diagnosis. The futility 'of attempting to base a diagnosis of a complex
systemic disease wholly upon laboratory data is fully realized. Therefore,
correlation of the history, clinical symptoms 'and laboratory findings is
emphasized.
My thanks are extended to the many persons who have contributed to
this edition. Words of encouragement and suggestions for improvements
have come from teachers in other schools -and from practicing veteri~

v

VI
. PREFACE

narians in various parts of the country. Dr: E. A. Benbrook's careful
perusal of the first edition is particularly appreciated. His criticisms
have improved the clarity of the text. Dr. Martin Kaplan's suggestions
for new material have been gratefully received and a number have been
included in this edition. Acknowledgment is made in the text to those
persons who have so generously contributed photographs and tabular
data.
I again wish to express my thanks to Dr. G. A. Dick for his encourage-
ment and to Dr. E. L. Stubbs for his help and advice in the field of clinical
pathology and for giving me access to his files for bibliographic material.
I also wish to acknowledge the contribution of Dr. H. M. Martin in lend-
ing the material from which the drawings of the mange mites and ana-
plasmata were made, and of Dr. H. W. Schoening in supplying the
material from which the drawings of piroplasmata were prepared. The
camera lucida used in the preparation of the majority of the original
drawings was graciously loaned by Dr. Josephine Deubler.
The efforts of the Comstock Publishing Company, Inc., in securing
the color plate of the Hotis Test for this edition, and its generally enthu-
siastic and co-operative policy are much appreciated.
The many line drawings and the water-color drawing of cattle blood
were done by my wife from specimens secured, in most instances, in the
laboratory. Several line drawings were added for this edition. Her work
on the manuscript has made this edition possible.

Philadelphia
July, 1945

D. L. C.

CONTENTS

Preface

I
MICROSCOPIC TECHNIQUE FOR CLINICAL P ATH-
OLOGIC EXAMINAdIONS

II HOW TO COLLECT, PACK AND SHIP §PECIMENS
FOR LABORATORY DIAGNOSIS

Why specimens spoil; Preservation of specimens; Handling of
specimens f~r specIfic dIseases or exammations

III PARASITOLOGIC EXAMINATIONS

Fecal-examinatIOn techniques; Nematode diagnosis; Tre~atode
diagnosIs; Lungworm dIagnosis; Ova charts for horses, dogs,
cats, foxes, swine, domestIc ruminants and poultry; Protozoan
diseases and dIagnostic tables for chickens, turkeys, sheep and
goats, hogs, dogs and cats, and cattle; Microfilaria examination

IV
URINE EXAMINATIONS

Physical examinatIOns; Chemical examinations; Field urinaly-
sis; MicroscopIc examination of sedIment

V INTERPRETATION OF URINARY FINDINGS

VI
HEMATOLOGY

Cells in Circulating blood; Collection of blood; Blood smears;
Study of the stamed smear; Avian blood; Differential count of
leukocytes; Hemoglobtn determination; Leukocyte enumera-
tion; Erythrocyte el1l1meration; Red-cell volume determination;
Reticulocyte enumeration; IcterUS-Index determination; Sedi-
mentation-rate determinatton;
B1ood-compatibihty
testing;
Care of equipment; Normal ranges for blood cellular elements;
Normal range of chem1lcal constituents

VII INTERPRETATION OF' HEMATOLOGIC FINDINGS

Hemoconcentration; Anemia; Leukocytosis; Leukopenia; Leu-
kemia; VariatIOns in blood plasma; SedimentatIOn rates;
Compatibility testing
vii

v

6

19

86

120

CONTf,NTS

VIII DIAGNOSTIC METHODS IN BACTERIAL DISEASE

Microscopic mspection; Culture; Serol~glc tests; Ammal mocu-
latlOn; Diagnostic tables for bacterial dlsea~e in cattie, horses,
hogs, sheep, domestic poultry, dogs, gumea pigS, mice and rat;,
and rabbtts
' j

IX
DIAGNOSIS OF BACTERIAL DISEASES

ActinomycosIs and actmobacdlo~;ts; Fusospirochetosis; lohne's
(iJsease; LeptospJrOSIS; Mastitis; SpeClfic cystitis and pyelo-
nephritis
.

X DIAGNOSIS OF MYCOTIC DISEASES

AspergillosIs; Favus; Rmgworm; Thrush

XI
DIAGNOSIS OF PROTOZOAN BLOOD DISEASES

Anaplasmosis; Bird malaria; Bovme piroplasmosIs; Canine
piroplasmosis; Leukocytozoon infectIOns
r

XII
DrAGNOSIS OF PROTOZOAN GENITAL iNFECTION

Rovine trichomoniasis

XIII DIAGNOSTIC METHODS IN VIRUS DISEASES

Hog cholera; Camne distemper; Equll1e encephalitis; Fox
encephalItIs; Infectious laryngotracheitIs; Panleukopenia of
cats, Bird pox; Pseudorabies; PSIttacosIs; Rabies; Equlllc vIrus
abortIOn; VIrus pneumoma of cats

XIV
FERTILITY EXAMINATIONS

Semen examInations; Diagnosis of pregnancy by laboratory
methods

XV
POULTRY AND GAME-BIRD AUTOPSY

XVI
FORMULAS AND TECHNIQUES

Solutions; StallB ancl techniques

INDEX

145

186

206

210

222

249

MANUAL OF Veterinary

Clinical Pathology

CHAPTER I

MICROSCOPIC TECHNIQUE

FOR CLINICAL P A THOLOG IC

EXAMIN A TIONS

T
HE MICROSCOPE is the most important single piece of equipment
used in laboratory diagnosis. Like a fine camera, however, it offers
so many possibilities of adjustment that it is often grossly mishandled
by the uninitiated. No microscope is better than its operatoL Therefore,
before beginning the study of Clinical Pathology, one should become
acquainted with and apply assiduously the principles of microscopy. For
detailed and complete information consult a textbook on the use of the
microscope.

USE OF THE MICROSCOPE

Study of the Unstained Smear
This procedure is used frequently. Most parasitologic, urinary-sediment
and unstained blood-cell examinations, such as blood counts, fall in this
class. The cardinal rule is: Work with the least light and the lowest power
consistent with good visibility.
For best illumination get light from a bright source, reflected straight
up the tube by means of the copcave mirror (central illumination). Ad-
just it by "stopping" it down by closing the iris diaphragm to its mini·
mum, then opening the aperturf until the best visibility is obtained. Use
the low-power objective (16 mm.) to locate the field and the objects for
study. Then, if it is required for identification, rotate the high dry ob-
jective (4 mm.) into place.
For clinical laboratory work, one should have-a high dry objective of

VETERINARY CLINICAL PATHOLOGY

sufficient working distance. Select a 4 mm·. lens with the proper N.A.
such as:

Bausch and Lomb
Spencer Len~ Co.
LeItz

4 mm. N.A. 0.65
(not 0.85)
4 mm. N.A. 0.66
(not 0.85)
No.6 l.g.
(not No.6)

Any of these lenses will give sufficient working distance for blood-cell
counts and examination of urinary seoiments and fecal smears.
Never focus down with the fine adjustment. Be especially careful to
prevent contamination of the high-power objective by immersion in the
Slnear.
Occasionally it is helpful to lower the condenser below the position of
maximum or critical illumination for some special purpose. This act is
much abused, however, since the condenser IS often lowered Inerely to
reduce the illumination, with consequent loss of definition. Instead, re-
duce the aperture of the iris diaphragm' to dim the light.
When third-dimensional vision is desired, as for the determination of
the cylindrical appearance of tube casts, oblique illumination can be ob-
tained by swinging the mirror to the side. This reflects the light at an
angle to the axis of the tube.
Almost constant adjustment of both the illummation and the focus
is required for maximum effect in the study of any fresh smear.

Study of the Stained Smear
Examinations of stained preparations, such as blood films, bacterial
smears and tissue sections, comprise this class. Bright lighting must be
used to bring out the color needed for differentiation. Central lighting
is best. With many preparations it is again desirable to use the 16 mm.
objective for the location of the field. This is particularly. true for
inclusion-body'studies of rabies or canine distemper where the proper
cells must be located before more detailed studies are attempted.
The plane mirror should be used when daylight or a special micro-
scopist's light is employed. The rays of such light are parallel. But when
divergent rays are used, as from an ordinary light bulb, the concave mir-,
ror should bc turned up. When examining tissue sections with the low
. or high dry objective, throw the condenser out of focus by lowering it
below the critical point or_ by using_thc_..Eoncave mirror and reducing the
iris diaphragm. Use critical illumination for- oilcimmersion work.

MICROSCOPIC TECHNIQUE
3

Study of the Dark-Field Preparation
Dark-field study is relatively new in veterinary medicine. It has been
emphasized recently because of the increasing importance of spirochetes
in canine practice. We now realize that Leptospira and the several mouth
Treponema organisms are damaging parasites and that dark-held exam-
inations can be employed as an aid in their diagnosis.
Dark-field microscopy also can be used' for special studies of organisms
that are visible by ordinary means. Thus, motility examinations of such
organisms as the coliforms and Salmonella species are easily performed,
and the flagella of bacteria and the processes of the ciliate and flagellate
protozoa are made plainly visible by this method.
A different principle of illumination is used. Here reflected light in-
stead of the usual refracted light is utilized. Objects in the field appear
as brightly lighted bodies against a black background in the same way
that the planets and the moon are visible in the darkened sky. Minute
objects beyond the limit of visibility with ordinary lighting become
plain'ly visible when reflected, oblique light is used. Most bacteria can be
seen at 100 X, and motility examinations of such organisms as the
coliforms may readily be made at 400 X. Many minute objects, visible as
reflected points of light and exhibiting active Brownian movement, are
seen in all dark-held preparations and should not be confused with
bacteria.

A,fechamcs of Dark-FIeld MIcroscopy. A speCial condenser must be used for maxi-
mum effect. The "paraboloid" or "cardIOid" condenser IS substituted for the ordi-
nary Abbe c~ndenser III the substage and is adjusted by lateral setscrews until it

IS centered With the aXIs of the tube. l\10st condensers have an etched ring or other
deVICe in the center of the opaque central area on which the low-power objective is
focused for alignment. A drop 6f fluid (Immersion od, glycenn or water) is then
placed on top of the conden$er, and it is "racked" up until it is in contact with the
slIde containing the preparation to be viewed. The adjustment of the lamp and
mirror should concentrate the LIght IIltensely III the flUid. The preparation is now
ready to be studied by low, high dry or ad-ImmersIOn techniques in the usual
manner.
The height of the condenser should be adjusted until the minute objects (vis!ble
in all preparations by dark Eeld) are receiving maximum light. For Oil-immersion
studIes a "funnel" stop with a ,small aperture must be placed 1Il the obJcctlvc un·
less the lens IS equipped With a diaphragm. This small opening prevents distorted
rays of hght from ~ntenng the objective and blurring the VisibiLIty of the field.
The preparation of the smear for study is slmilar_ to that of any fresh specimen.

4
VETERINARY CLINICAL PATHOLOGY

Clean slides and cover glasses must be used. (See Pusospirochetosis, p. 188, and
Canine Leptospirosis, p. 191.)

ILLUMINA YION FOR THE MICROSCOPE

In microscopy the source of illumination is fully as important as the
microscope. One should consider the two as interdependent. When pur-
chasing a microscope, therefore, or seeking a place to install one in an
office or laboratory, give proper thought to the light source. There are
many types of lamps. A few are excellent, some adequate and many
inferior.
Daylight may be used providing a north window is selected with a

view of the sky uninterrupted by either buildings or trees. No other ex-
posure can be used because the direct ra ys of the sun are sure to be present
sometime during the day, making microscopy practically impossible.
Daylight has many limitations. It is never bright enough for good oi1-
immersion work when a binocular microscope is used or for dark-field

Fig. I. Inexpensive microscope light suitable for
monocular or binocular microscopes. (Courtesy
Bausch & Lomb Optical Co.)

illumination. And, if any type
of compound instrument is to
be used on cloudy days or after
dark, some source of artificial
light is demanded.
Artificial light is the only
source of constant and uniform
lighting of the proper inten-
sity. Some lamps are rather ex-
pensive,
others inexpensive.
Excellent lamps may be im-
provised from very inexpensive
materials. The small substage
light is satisfactory for mo-
nocular microscopes. I t rna y be
placed directly beneath the

stage or in front of the microscope when the mirror is used. It will not,
however, furnish light intense enough for oil-immersion studies with
binocular instruments or for dark-field work. The small swivel type of

MICROSCOPIC TECHNIQUE
5

light is economical and adequate for binocular microscopes when fitted
with a 60-watt daylight bulb. Several styles are available.

The many types of more expen-
sive lamps on the market that use
lenses or water flasks to produce
parallel rays are excellent. (For oc-
casional work a 40- or 60-watt day-
light bulb may be used alone.)
One can improvise such a lamp eco-
notnically by means of an opaque
Fig. 2. Diagram of improvised lamp using

box containing the socket and bulb
water flask as light filter.

and having a 4- or 4 Yz-inch circular hole in one end. A 6-inch globe, filled
with dilute copper sulphate solution, when htted into the aperture, will
produce blue-tinted light with parallel rays. A similar effect may be
achieved by substituting a frosted or ground-glass plate for the flask and
using a blue-tinted ,bulb as the source of light.

REFERENCES
Gage, S. H. 1941. Th e Microscope. Comstock Publishing Co., Inc., Ithaca, N.Y.